Conformational rearrangements in 2nd voltage sensor domain switch PIP2- and voltage-gating modes in two-pore channels

Two-pore channels (TPCs) are activated by PIP2 binding to domain I and/or by voltage-sensing in domain II (DII). Little is known about how these two stimuli are integrated and how each TPC subtype achieves its unique preference. Here, we show that the distinct conformations of DII-S4 in the voltage-sensor domain determine the two gating modes. DII-S4 takes an intermediate conformation, and forced stabilization in this conformation was found to give or maintain a high PIP2-dependence in primarily voltage-dependent TPC3 or in PIP2-gated and non-voltage-dependent TPC2, respectively. We also found in TPC2 that a tricyclic antidepressant desipramine induces the DII-S4 based voltage-dependence and that a flavonoid naringenin biases the mode preference from PIP2-gating to desipramine-induced voltage-gating. Taken together, our study on TPCs revealed an unprecedented mode-switching mechanism involving conformational changes in DII-S4. This will pave the way for drug development by targeting specific gating modes of TPCs. ### Competing Interest Statement The authors have declared no competing interest.